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INTRODUCTION: Osteoblasts (OBs) and osteoclasts (OCs) interaction have an important role in bone remodeling [1]. Thus, coculture with both bone cells (OB-OC) is needed to achieve a better understanding of the material-cell interaction and the impact on Mg degradation for its potential use as a biomedical material. The aim of this study was to analyze the surface morphology and dynamic changes in the composition of the corrosion layer formed on Mg influenced by direct monoculture and coculture models.
METHODS: High purity Mg (99.94%) disc-shaped samples (diameter 9 mm, thickness 1.5 mm) were used in this study. The specimens were grinded, polished, and finally sterilized by gamma irradiation at a dose of 33.5 kGy.
α-MEM (Minimum Essential Medium) with 10% FBS (Fetal Bovine Serum) and 1% P/S (Penicillin/Streptomycin) was used as cell culture and immersion medium. Human primary OBs and OCs were pre-differentiated separately for 21 days (from HUCPV and PBMC *, respectively) and seeded on materials as described in Ref [2] in the monoculture, and coculture model (OB 1: 2 OC ratio). Cells also were seeded on glass (herein, employed as controls). Changes in adhesion were evaluated by actin cytoskeleton staining after 7 and 14 days. Surface topography and cell morphology were analyzed on samples prepared by Critical Point Drying (CPD) by SEM (SU-8000, Hitachi). Cross-sectioning, imaging, and evaluation of distributed elements composing the degradation interface were analyzed by FIB/SEM/EDX (NB-5000 dual-beam, Hitachi) from samples prepared by CPD.
RESULTS: Cell spreading, the proliferation of differentiated OBs, and cell attachment of OCs were observed after 7- and 14-days cell culture on Mg. OBs extensively grew as elongated cells whereas rounded adhered multinucleated OCs showed their typical dorsal villus-like processes and adhesion to the Mg surface. After 14 days, the corrosion layer formed under OCs had slightly higher and uniform distribution weight percent (wt.%) oxygen content. Similarly, Phosphorus (P) and Calcium (Ca) were slightly higher in wt.% content and homogeneously distributed in the corrosion layer formed beneath the OB-OC coculture.
CONCLUSIONS: The results showed the successful cell attachment and the vitality of differentiated OBs and OCs on Mg surface in direct cell monoculture and coculture models after 14 days. A slightly distinct corrosion product was pointed out beneath OB-OC coculture with slightly homogenously increased wt.% of P and Ca compared to monoculture models after 14 days.
REFERENCES: [1] Borciani, G. et al., Acta Biomater. 108, 22-45 (2020). [2] Agha, N. et al., PLoS ONE, 11, 1-20 (2016).
ACKNOWLEDGEMENTS: The research leading to these results has received funding from the European Training Network within the framework of Horizon 2020 Marie Skłodowska-Curie Action (MSCA) No. 811226.
- HUCPV: Human Umbilical Cord Perivascular Cells
** PBMC: Peripheral Blood Mononucleated Cells
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